All current mouse models of human liver allow only HBV replication in the absence of a functional immune system. Those models are not useful to studying host immune responses, their contributions to HBV pathogenesis and potential immunotherapeutic approaches. The Rag2-gC double knock (DKO) mouse lacks T/B and NK cells, and allows development of a functional human immune system with human Hematopoietic Stem Cell (DKO-hu HSC mice). Normal human T, B, and dendritic cells are present in lymphoid tissues such as thymus, spleen, peripheral blood (PB) and lymph nodes (LN). Recently, we have also co-engrafted human hepatocytes with human immune system in the DKO mouse transplanted with human hepatocyte progenitor cells and CD34+ HSC cells (DKO-hu HSC/Hep mice). I propose to optimize the DKO-hu HSC/Hep mouse to increase human hepatocyte engraftment by selectively depleting murine hepatocytes and promoting human hepatocytes in the DKO-hu HSC/Hep mouse. We will also study HBV infection, immuno-pathogenesis, and development of hepatocellular carcinoma (HCC) in this model. 1) To optimize the DKO-hu HSC/Hep model. a. Improving DKO-hu-HSC/Hep mice with preferential murine hepatocyte depletion in vivo. I propose to deplete murine hepatocytes in AFC8/DKO hu HSC/Hep mice (transgenic DKO mice with FKBP-caspase8 fusion gene under control of the albumin promoter, 14, 27) with FKBP dimerizer AP20187. b. Boosting human hepatocyte cell growth with anti-human c-Met mAb. To improve human hepatocyte growth in DO-hu-HSC/Hep mice, we will use an agonistic antibody against human c-Met (c-Met mAb, mouse lgG1) that activates human but not murine c-Met(46). 2) To study HBV infection and pathogenesis in DKO-hu HSC/Hep mice. a. HBV infection and immuno-responses in the DKO-hu HSC/Hep model. We will establish HBV infection kinetics, immuno-responses and pathogenesis in the current DKO-hu HSC/Hep model and in the improved models from SA1. b. the effects of ethanol on HBV infection, immuno-pathogenesis and development of hepatocellular carcinoma (HCC) in DKO-hu HSC/Hep model. DKO-hu HSC/Hep mice infected with HBV will be treated with acute and chronic alcohol consumption. HBV replication, immune responses, pathogenesis and cancer development in DKO-hu HSC/Hep mice will be investigated. Public Health Relevance: The long-term goals of this project are to develop a relevant mouse model to investigate the immuno- pathogenesis of HBV infection, the mechanism of HCC development, and to model immune-based therapy. Specifically, we will develop the DKO-hu HSC/Hep model for studying HBV infection and immuno- pathogenesis in the absence or presence of alcohol-induced liver insults. These studies will establish the foundation for future study and shed light on novel therapeutic strategies for controlling HBV diseases.